Oxidoreductase is a kind of enzyme being most widely used after hydrolase, most of the oxidation-reduction reactions catalyzed by it needs NAD(P)H/NAD(P)+ to provide reduced hydrogen or to receive hydrogen, but high price of NAD(P)H/NAD(P)+ restricts use of the oxidoreductase in industrial production, requiring conducting in situ regeneration of coenzyme to reduce the cost. At present, the coenzyme regeneration includes five methods of whole-cell method, enzyme method, electrochemical method, photochemical method and chemical method. For the whole-cell method, its cost is low, but product separation is complicated, the method is comparatively primitive. The enzyme method includes two methods of substrate coupling method and enzyme coupling method, wherein the substrate coupling method is the same enzyme catalyze both of two different substrates towards reverse directions, meanwhile achieving conversion of main substrate and co-substrate, forming a coenzyme regeneration circulation, co-substrate and main substrate compete an active center of the enzyme, a part of the enzyme activity is used in regeneration of coenzyme, thus the enzyme activity used in catalyzing the main substrate is reduced; in the enzyme coupling method, the oxidization reaction enzyme system and the reduction reaction enzyme system works in one system in parallel, production of the enzyme catalyzes substrate conversion and consumes coenzyme, regeneration of the enzyme catalyzes regeneration of the coenzyme, this method requires that the substrates in two enzyme systems must be independent to each other, to avoid mutual competition to active center of the same enzyme, and optimum reaction condition for these two enzymes are often inconsistent. For the eletrochemical method, the cost of electricity is low, there is no stoichiometric regeneration reagent, but the efficiency is low, often requiring participation of a medium, and this method is incompatible to many biochemical catalytic systems, its selectivity is poor. The chemical method uses a special catalyst to transfer oxid-reduction equivalent of the oxidant and reductant to a redox cofactor NAD(P)H/NAD(P)+ thereby forming a cofactor regeneration circulation, the commonly used chemical catalyst is a noble metal organic complex (Rh/Ru/Ir), its price is high and center metal may fall off, the center metal often coordinates with amino acid residue of the enzyme, making the metal catalyst and the enzyme both losing activity. The photochemical regeneration method is often limited by deficiency of efficient photosensitizer, thus generally its efficiency is low.
The chemical method currently used in regeneration of the oxidized cofactor NAD(P)+ often uses two kinds of regeneration catalyst for riboflavin mononucleotide (FMN) [J. Am. Chem. Soc. 1982, 104, 4659-4665] and iron (III) porphyrin [Angew. Chem. Int. Ed. 2011, 50, 2397-2400]. When the FMN is used in the regeneration, it is required to use overdose [J. Am. Chem. Soc. 1982, 104, 4659-4665] or under radiation of a special light source [Chem Cat Chem 2011, 3, 338-342] to conduct catalytic regeneration. The iron porphyrin belongs to metal organic complex, its regeneration efficiency is not high, and tends to coordinate with the enzyme, resulting in deactivation.